1. Field of the Invention
The present invention relates generally to methods for analyzing biological samples, and more particularly, to the assessment of clinical biological sectioned samples using computerized digitizing techniques.
2. Description of the Relevant Art
The incidence of non-melanoma skin cancers is increasing dramatically throughout the United States. The vast majority of these cancers develop in areas of severe sun damage, particularly within or contiguous to areas of solar keratoses, but it has not been possible thus far to predict who will develop these potentially dangerous skin cancers. Although the mortality rate for these skin cancers is low, their treatment is associated with considerable morbidity and significant medical care costs.
Efforts to characterize the histopathological and karyometric changes induced by solar keratosis face difficulties in defining a standard. First, there is the variety of skin types. At best, "skin type" constitutes a fuzzy categorization. There is the difficulty of defining exposure, its nature and total duration for a given individual; this source of variation requires that comparisons be made between a visually apparent region of solar keratosis and an adjacent, seemingly unaffected skin area. Such an area, in all likelihood, would have received a comparable dose of solar irradiation. Well-defined samples of entirely unexposed skin, desirable for comparison with the changes induced by solar radiation, are usually not available. One option is a comparison to skin biopsies taken from remote, minimally exposed body sites. At this time there exists no database that characterizes the nuclear chromatin features of skin samples of such an origin.
Similar problems arise in attempts to define the efficacy of chemopreventive treatment of solar keratosis lesions. One cannot take a second biopsy from the original lesion since epidermal change and dermal fibrosis may introduce additional variability. If a different solar keratosis lesion is biopsied to assess treatment efficacy, one may bias the sampling because the selected site constitutes a lesion that most likely persisted throughout treatment.
The same general problems arise when attempting to detect and analyze other types of lesions in tissues other than the skin, e.g., breast lesions, prostatic lesions, endometrial legions, esophogeal lesions, etc. Currently known methods of analyzing clinical samples of such tissues tend to be qualitative in nature, and are often subjective rather than objective. Moreover, currently known methods of analyzing such clinical samples do not detect changes in the nature of such lesions except after relatively long periods of time. Evaluation of premalignant lesions is typically made by a visual assessment and is expressed in terms of categories of lesions, such as "hyperplasia", or "severe adenoma", or as a grade, such as grade II or grade IV, or as "low grade" and "high grade". Such linguistic labels are vague and difficult to reproduce consistently between observers, or even by the same observer. As measures to assess the efficacy of a chemopreventive agent, they lack precision and the ability to detect a small, but significant change. Nevertheless, the existing diagnostic terms, fuzzy and vague as they are, do represent the current state of knowledge and ultimately the standard from which the clinical effect of treatment by chemopreventive agents has been assessed.
The study of sectioned images of lesions is generally known. For example, in the technical article by Bostwick and Brawer entitled "Prostatic Intra-Epithelial Neoplasia and Early Invasion in Prostate Cancer", Cancer 59:788-794, 1987, the authors describe the process of marking prostatic basal cell layers using a monoclonal antibody to certain keratin proteins. Disruption of the basal layer was used to detect early invasion in prostate cancer. Among the characteristics studied by Bostwick and Brawer were crowding and multilayering of cells, variation in nuclear size and the prominence of nucleoli. In most instances, subjective inspection of the histological appearance of sectioned and stained material is carried out by experienced pathologists, but despite the experience of such individuals, the process remains subjective and qualitative.
In the paper by Bacus, Bacus, Stoner, Moon, Kelloff, and Boone entitled "Quantitation of preinvasive neoplastic progression in animal models of chemical carcinogenesis", J Cell Biochem Suppl, 1997; 28-29:21-38, the authors describe the use of image analysis through the use of high-resolution tiled images of complete tissue sections to quantitate cellular and tissue changes associated with early, preinvasive neoplasia. The image analysis was applied to histological sections of tissue that had been sectioned and stained. The authors describe a histological grading system, or scale, expressed in normal deviate units of multiple and different morphometric descriptors. However, those skilled in the art have as yet failed to provide a method of quantitatively expressing how far a tissue has progressed from normal to malignant disease. Likewise, those skilled in the art have not provided a method for reliably detecting changes in the status of tissue under study in a relatively prompt manner. If reliable detection and quantitative assessment of such changes could be performed relatively quickly, then the efficacy of chemopreventive drugs or treatments could be evaluated much more quickly, and promising drugs and/or treatments could be brought into use in shorter periods of time.
Accordingly, it is an object of the present invention to provide a method for quantitatively measuring the progression of a lesion along a progression curve ranging between normal tissue and malignant disease.
A further object of the present invention is to provide a method of objectively assessing and grading sampled tissue to assign a progression index value to a lesion.
A still further object of the present invention is to quantitatively detect and assess changes in tissue over a relatively short period of time.
Yet another object of the present invention is to rapidly and quantitatively determine the efficacy of a chemopreventive drug or treatment in combating progression of tissue from its normal state toward malignant disease.
These and other objects of the present invention will become more apparent to those skilled in the art as the present description proceeds.